REPRODUCING THE NATURAL EVOLUTION OF PROTEIN STRUCTURAL FEATURES WITHTHE SELECTIVELY INFECTIVE PHAGE (SIP) TECHNOLOGY - THE KINK IN THE FIRST STRAND OF ANTIBODY KAPPA-DOMAINS

The beta-sandwich structure of immunoglobulin variable domains is char acterized by a typical kink in the first strand, which allows the firs t part of the strand to hydrogen bond to the outer beta-sheet (away fr om the V-H-V-L interface) and the second part to the inner beta-sheet. This kink differs in length and sequence between the V-kappa, V-lambd a and V-H domains and yet is involved in several almost perfectly cons erved interactions with framework residues. We have used the selective ly infective phage (SIP) system to select the optimal kink region from several defined libraries, using an anti-hemagglutinin single-chain F v (scFv) fragment as a model system. Both for the kink with the V-kapp a domain length and that with the V-lambda length, a sequence distribu tion was selected that coincides remarkably well with the sequence dis tribution of natural antibodies. The selected scFv fragments were puri fied and characterized, and thermodynamic stability was found to be th e prime factor responsible for selection. These data show that the SIP technology can be used for optimizing protein structural features by evolutionary approaches. (C) 1998 Academic Press.